Next-Generation Sequencing of Endobronchial Ultrasound Transbronchial Needle Aspiration Specimens in Lung Cancer

Fielding, David; Dalley, Andrew J.; Bashirzadeh, Farzad; Singh, Mahendra; Nandakumar, Lakshmy; Reed, Amy E. McCart; Black, Debra; Kazakoff, Stephen H.; Nones, Kátia; Pearson, John V.; Waddell, Nicola; Lakhani, Sunil R.; Simpson, Peter T.

doi:10.1164/rccm.201609-1935le

Cited by 14 publications

(13 citation statements)

References 10 publications

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“…Of the malignancies, primary lung cancer was diagnosed in 64 cases: adenocarcinoma (43 cases), squamous cell carcinoma (10 cases), and small-cell lung cancer (11 cases). Data relating to the NGS of cell block material from this study cohort are detailed elsewhere [31]. Briefly, there were 38 males and 29 females with a mean age of 67 ± 10 years.…”

Section: Resultsmentioning

confidence: 99%

“…For discordant calls where variants were called only in one DNA source, investigation of lower levels of evidence was carried out, using pileup at the variant position and manual review using Integrative Genomics Viewer. DNA sequencing of cell block samples has been described previously [31] and was reanalysed here for comparison to the sequence data obtained from the smears.…”

Section: Methodsmentioning

confidence: 99%

“…We previously reported a prospective cohort of 76 cases who had NGS performed using DNA from EBUS- TBNA-derived FFPE cell blocks [31]. All these cases also had ROSE smears made, and so here we report on an assessment of the cellularity (abundance) of the slides, their DNA yields, and their performance in amplicon-based gene panel NGS.…”

Section: Introductionmentioning

confidence: 99%

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Diff-Quik Cytology Smears from Endobronchial Ultrasound Transbronchial Needle Aspiration Lymph Node Specimens as a Source of DNA for Next-Generation Sequencing Instead of Cell Blocks

et al. 2019

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Background: Next-generation sequencing (NGS) in lung cancer specimens from endobronchial ultrasound transbronchial needle aspiration (EBUS-TBNA) is usually performed on formalin-fixed paraffin-embedded cell block material. Objectives: Since DNA can be damaged by this process, we investigated the potential of using DNA extracted from Diff-Quik cytology smears made for rapid on-site evaluation during EBUS-TBNA. Methods: In a prospective study, 67 patients undergoing diagnostic EBUS-TBNA were analysed. We compared cell blocks and smears for DNA yields and sequencing (TruSeq Amplicon Cancer Panel) outcomes. Smears were also evaluated for tumour cell fraction and overall cellularity (cell count). Results: Primary lung cancer was diagnosed in 64 patients and metastatic malignancy in 3 patients. The DNA yield from smears was significantly higher than that obtained from matched cell blocks (mean 1,740 vs. 434 ng; p = 0.001). For 33 cases with matched smears and cell blocks the mutation profiles were similar. Smears with abundant malignant cells (using a cut-off of > 25% tumour cell fraction and > 1,000 cells) accurately predicted high (> 50 ng) DNA yield and therefore success in triaging samples to sequencing. In terms of tissue workflow, using only smears as source DNA for sequencing was an improvement in the use of only cell blocks (54/67 [80.6%] vs. 41/67 [61.2%]); however, the use of cell blocks when smears were not available or did not yield sufficient DNA further improved the success rate to 62/67 (92.5%) cases. Conclusion: We recommend smears in laboratory workflows as the primary source of DNA for NGS following an EBUS procedure.

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Section: Resultsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

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Diff-Quik Cytology Smears from Endobronchial Ultrasound Transbronchial Needle Aspiration Lymph Node Specimens as a Source of DNA for Next-Generation Sequencing Instead of Cell Blocks

et al. 2019

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“…29,30 Identifying areas for improvement can be difficult, because several pre-analytical variables are involved, including in acquiring and processing a specimen. 34 Second, our results showed that of the cases that failed MT, 55% had ALK and PD-L1 (currently for histology specimens but being evaluated on cytology specimens 54 ) was approved and may become routine. Though noncytology factors (e.g., size of lymph node) may also influence MT, the results of the study would not be invalidated given the statistically significant differences.…”

Section: Slide Reviewmentioning

confidence: 78%

“…31 Some malignant nodes have high neoplastic content whereas others have greater stroma and vessels interspersed; the former may have higher cellular yield than the latter. 34 Thus, factors other than size may also impact MT.…”

Section: Discussionmentioning

confidence: 99%